Genomics of the human nucleolus – implications for cancer and aging
Supervisors: Prof. Brian McStay (NUIG), Prof. Cathal Seoighe (NUIG)
The primary role of the nucleolus is ribosome biogenesis but, more recently, its importance in the control of cell growth and proliferation, stress sensing and viral replication have become more fully appreciated. Nucleoli play a key role in the progression of many human diseases, including cancer. They form around arrays of ribosomal gene repeats termed the nucleolar organiser regions (NORs). A decade after the publication of the sequence of the human genome, the short arms of the acrocentric chromosomes (HSA13, 14, 15, 21 and 22), containing these repeats, are absent from the assembled human genome sequence. Consequently the precise organization of rDNA repeats within NORs and their chromosomal context remains poorly understood. Recent research using molecular combing techniques suggests that as much as 30% of rDNA gene repeats may be inverted or rearranged. Such alternative rDNA repeat configurations are hypothesized to have important implications in cancer and possibly aging.
At NUI Galway we have developed protocols for preparing DNA from nucleoli isolated from human cells. This DNA is highly enriched for acrocentric chromosome short arm DNA, including NORs, as determined by back painting onto metaphase chromosomes. Molecular combing of nucleolar DNA confirms that a large proportion of rDNA repeats within NORs are rearranged. We have used nucleolar DNA as a template in next generation (454) sequencing and have obtained short-read sequence datasets, with approximately 10% of sequence reads mapping to ribosomal RNA genes. The objectives of this project will be to assemble and makes sense of these sequence reads. Assembled reads will then be used to investigate the structure of rDNA repeats as well as the sequences flanking the rDNA gene clusters. Coverage of the acrocentric chromosomes is sufficient to provide a high probability of confirming or refuting the existence of rearranged rDNA gene clusters with profound implications for the function of these key and poorly understood gene clusters. Having characterised the structure of rearranged rDNA repeats, a series of PCR primers will be developed to permit the analysis of rearranged rDNA in human diseases and aging.
References
Pederson T. 2010. The Nucleolus. Cold Spring Harb Perspect Biol.
McStay B, Grummt I. 2008. The epigenetics of rRNA genes: from molecular to chromosome biology. Annu Rev Cell Dev Biol 24: 131-157.
Boulon S, Westman BJ, Hutten S, Boisvert FM, Lamond AI. 2010. The nucleolus under stress. Mol Cell 40: 216-227.
• 4 year stipend of €18,000 per year, plus fees and travel expenses
• Budget for personal laptop/PC
For the application procedure please visit:
http://bioinfo-casl.ucd.ie/PhD/index.php?option=com_content&view=article&id=23&Itemid=33
Supervisors: Prof. Brian McStay (NUIG), Prof. Cathal Seoighe (NUIG)
The primary role of the nucleolus is ribosome biogenesis but, more recently, its importance in the control of cell growth and proliferation, stress sensing and viral replication have become more fully appreciated. Nucleoli play a key role in the progression of many human diseases, including cancer. They form around arrays of ribosomal gene repeats termed the nucleolar organiser regions (NORs). A decade after the publication of the sequence of the human genome, the short arms of the acrocentric chromosomes (HSA13, 14, 15, 21 and 22), containing these repeats, are absent from the assembled human genome sequence. Consequently the precise organization of rDNA repeats within NORs and their chromosomal context remains poorly understood. Recent research using molecular combing techniques suggests that as much as 30% of rDNA gene repeats may be inverted or rearranged. Such alternative rDNA repeat configurations are hypothesized to have important implications in cancer and possibly aging.
At NUI Galway we have developed protocols for preparing DNA from nucleoli isolated from human cells. This DNA is highly enriched for acrocentric chromosome short arm DNA, including NORs, as determined by back painting onto metaphase chromosomes. Molecular combing of nucleolar DNA confirms that a large proportion of rDNA repeats within NORs are rearranged. We have used nucleolar DNA as a template in next generation (454) sequencing and have obtained short-read sequence datasets, with approximately 10% of sequence reads mapping to ribosomal RNA genes. The objectives of this project will be to assemble and makes sense of these sequence reads. Assembled reads will then be used to investigate the structure of rDNA repeats as well as the sequences flanking the rDNA gene clusters. Coverage of the acrocentric chromosomes is sufficient to provide a high probability of confirming or refuting the existence of rearranged rDNA gene clusters with profound implications for the function of these key and poorly understood gene clusters. Having characterised the structure of rearranged rDNA repeats, a series of PCR primers will be developed to permit the analysis of rearranged rDNA in human diseases and aging.
References
Pederson T. 2010. The Nucleolus. Cold Spring Harb Perspect Biol.
McStay B, Grummt I. 2008. The epigenetics of rRNA genes: from molecular to chromosome biology. Annu Rev Cell Dev Biol 24: 131-157.
Boulon S, Westman BJ, Hutten S, Boisvert FM, Lamond AI. 2010. The nucleolus under stress. Mol Cell 40: 216-227.
EU STUDENTS ONLY
Each student receives:• 4 year stipend of €18,000 per year, plus fees and travel expenses
• Budget for personal laptop/PC
For the application procedure please visit:
http://bioinfo-casl.ucd.ie/PhD/index.php?option=com_content&view=article&id=23&Itemid=33